University of Exeter View Institution's Website 13 articles published in JoVE Neuroscience Millisecond Hydrogen/Deuterium-Exchange Mass Spectrometry for the Study of Alpha-Synuclein Structural Dynamics Under Physiological Conditions Neeleema Seetaloo1, Jonathan J. Phillips1 1Living Systems Institute, Department of Biosciences, University of Exeter The structural ensemble of monomeric alpha-synuclein affects its physiological function and physicochemical properties. The present protocol describes how to perform millisecond hydrogen/deuterium-exchange mass spectrometry and subsequent data analyses to determine conformational information on the monomer of this intrinsically disordered protein under physiological conditions. Bioengineering Biomolecular Imaging of Cellular Uptake of Nanoparticles using Multimodal Nonlinear Optical Microscopy Chun-Chin Wang*1, Jessica C. Mansfield*1, Nicholas Stone1, Julian Moger1 1Biomedical Physics, School of Physics and Astronomy, University of Exeter This article presents the integration of a spectral-focusing module and a dual-output pulse laser, enabling rapid hyperspectral imaging of gold nanoparticles and cancer cells. This work aims to demonstrate the details of multimodal nonlinear optical techniques on a standard laser scanning microscope. Chemistry Uncovering Hidden Dynamics of Natural Photonic Structures Using Holographic Imaging Marina Simovic-Pavlovic1,2, Maja C. Pagnacco3, Dusan Grujic1, Bojana Bokic1, Darko Vasiljevic1, Sébastien Mouchet4,5, Thierry Verbiest6, Branko Kolaric1,7 1Institute of Physics, Photonics Center, University of Belgrade, 2Faculty of Mechanical Engineering, University of Belgrade, 3Institute of Chemistry, Technology and Metallurgy, University of Belgrade, 4School of Physics, University of Exeter, 5Department of Physics & Namur Institute of Structured Matter (NISM), University of Namur, 6Chemistry Department, Laboratory of Molecular Imaging and Photonics, KULeuven, 7Université de Mons The paper is primarily focused on the combined power of optical (linear and nonlinear) and holographic methods used to reveal phenomena at the nanoscale. The results obtained from the biophotonic and oscillatory chemical reactions' studies are given as representative examples, highlighting holography's ability to reveal dynamics at a nanoscale. Biology Preparation of Rat Skeletal Muscle Homogenates for Nitrate and Nitrite Measurements Ji Won Park*1, Samantha M. Thomas*1, Lee J. Wylie2, Andrew M. Jones2, Anni Vanhatalo2, Alan N. Schechter1, Barbora Piknova1 1Molecular Medicine Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 2Sport and Health Sciences, College of Life and Environmental Sciences, St Luke’s Campus, University of Exeter We present protocols for three different methods for the homogenization of four different muscle groups of rat skeletal muscle tissue to measure and compare the levels of nitrate and nitrite. Furthermore, we compare different sample weights to investigate whether tissue sample size affects the results of homogenization. Immunology and Infection Field Postmortem Rabies Rapid Immunochromatographic Diagnostic Test for Resource-Limited Settings with Further Molecular Applications Stephanie Mauti1, Monique Léchenne2, Service Naïssengar3, Abdallah Traoré4, Vessaly Kallo5,6, Casimir Kouakou7, Emmanuel Couacy-Hymann7, Morgane Gourlaouen8, Céline Mbilo9,10, Pati Patient Pyana11, Enos Madaye3, Ibrahima Dicko4, Pascal Cozette1, Paola De Benedictis8, Hervé Bourhy1, Jakob Zinsstag9,10, Laurent Dacheux1 1Unit Lyssavirus Epidemiology and Neuropathology, National Reference Center for Rabies and WHO Collaborating Center for Reference and Research on Rabies, Institut Pasteur, 2Environment and Sustainability Institute, University of Exeter, Penryn Campus, 3Institut de Recherche en Elevage pour le Développement, 4Laboratoire Central Vétérinaire, 5 We present a complete protocol for postmortem diagnosis of animal rabies under field conditions using a rapid immunochromatographic diagnostic test (RIDT), from brain biopsy sampling to final interpretation. We also describe further applications using the device for molecular analysis and viral genotyping. Medicine Assessment of Kidney Function in Mouse Models of Glomerular Disease Megan Stevens1,2,3, Sebastian Oltean1,2,3 1Institute of Biomedical and Clinical Sciences, Medical School, University of Exeter, 2School of Physiology, Pharmacology and Neurosciences, University of Bristol, 3Bristol Renal, School of Clinical Sciences, University of Bristol This protocol describes a full kidney work-up that should be carried out in mouse models of glomerular disease. The methods allow for detailed functional, structural, and mechanistic analysis of glomerular function, which can be applied to all mouse models of glomerular disease. Bioengineering Creating a Structurally Realistic Finite Element Geometric Model of a Cardiomyocyte to Study the Role of Cellular Architecture in Cardiomyocyte Systems Biology Vijay Rajagopal1,2,3, Gregory Bass2,3, Shouryadipta Ghosh1,2,3, Hilary Hunt2,4, Cameron Walker5, Eric Hanssen6, Edmund Crampin2,3,4,7,8, Christian Soeller9 1Cell Structure and Mechanobiology Group, University of Melbourne, 2Systems Biology Laboratory, Melbourne School of Engineering, University of Melbourne, 3Department of Biomedical Engineering, University of Melbourne, 4School of Mathematics and Statistics, Faculty of Science, University of Melbourne, 5Department of Engineering Science, University of Auckland, 6Advanced Microscopy Facility, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, 7ARC Centre of Excellence in Convergent Bio-Nano Science and Technology, University of Melbourne, 8School of Medicine, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, 9Living Systems Institute, University of Exeter This protocol outlines a novel method to create a spatially detailed finite element model of the intracellular architecture of cardiomyocytes from electron microscopy and confocal microscopy images. The power of this spatially detailed model is demonstrated using case studies in calcium signaling and bioenergetics. Genetics Metagenomic Analysis of Silage Richard K. Tennant1, Christine M. Sambles1, Georgina E. Diffey1, Karen A. Moore1, John Love1 1Biosciences, University of Exeter Metagenomics was used to investigate the microbiome of silage cattle feed. Analysis was performed by shotgun sequencing. This approach was used to characterize the composition of the microbial community within the cattle feed. Bioengineering Preparation of Extracellular Matrix Protein Fibers for Brillouin Spectroscopy Ryan S. Edginton1, Sara Mattana2, Silvia Caponi2,3, Daniele Fioretto2, Ellen Green1, C. Peter Winlove1, Francesca Palombo1 1School of Physics and Astronomy, University of Exeter, 2Department of Physics and Geology, University of Perugia, 3Istituto Officina dei Materiali del CNR, Unità di Perugia We present a protocol for the application of Brillouin light scattering spectroscopy to elastin and trypsin-purified type I collagen fibers of the extracellular matrix to extract their full elastic properties. Biology The Infiltration-centrifugation Technique for Extraction of Apoplastic Fluid from Plant Leaves Using Phaseolus vulgaris as an Example Brendan M. O'Leary1, Arantza Rico2, Sarah McCraw1, Helen N. Fones3, Gail M. Preston1 1Department of Plant Sciences, University of Oxford, 2School of Education of Vitoria-Gasteiz, University of the Basque Country (UPV/EHU), 3Biosciences, University of Exeter This protocol details the optimized extraction of apoplast washing fluid from plant leaves, using French bean plants (Phaseolus vulgaris) as a model example. Biology Determination of Protein-ligand Interactions Using Differential Scanning Fluorimetry Mirella Vivoli1, Halina R. Novak1, Jennifer A. Littlechild1, Nicholas J. Harmer1 1Department of Biosciences, University of Exeter Differential scanning fluorimetry is a widely used method for screening libraries of small molecules for interactions with proteins. Here, we present a straightforward method to extend these analyses to provide an estimate of the dissociation constant between a small molecule and its protein partner. Immunology and Infection Detection of Invasive Pulmonary Aspergillosis in Haematological Malignancy Patients by using Lateral-flow Technology Christopher Thornton1, Gemma Johnson2, Samir Agrawal3 1Biosciences, University of Exeter, 2BICMS, Queen Mary University of London, 3St. Bartholomew's Hospital and The London NHS Trust A rapid and accurate point-of-care test for invasive pulmonary aspergillosis is presented. It takes advantage of lateral-flow technology using a specific monoclonal antibody that binds to an Aspergillus antigen secreted during pulmonary infections. The assay is compatible with serum and brochoalveolar lavage and represents a novel adjunct test for disease diagnosis. Biology A Simple Method for Imaging Arabidopsis Leaves Using Perfluorodecalin as an Infiltrative Imaging Medium George R. Littlejohn1, John Love1 1Biosciences, College of Life and Environmental Sciences, The University of Exeter We describe the use of perfluorodecalin as an infiltrative mounting medium. This is a simple method for improving depth of imaging in Arabidopsis thaliana leaf tissue with minimal physiological impact.